High density continuous production of murine pluripotent cells in an acoustic perfused bioreactor at different oxygen concentrations.

Ricardo P Baptista, David A Fluri, Peter W Zandstra

Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada.

Biotechnology and bioengineering 2013 Feb

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Strategies for the production of pluripotent stem cells (PSCs) rely on serially dissociated adherent or aggregate-based culture, consequently limiting robust scale-up of cell production, on-line control and optimization of culture conditions. We recently developed a method that enables continuous (non-serially dissociated) suspension culture-mediated reprogramming to pluripotency. Herein, we use this method to demonstrate the scalable production of PSCs and early derivatives using acoustic filter technology to enable continuous oxygen-controlled perfusion culture. Cell densities of greater than 1 × 10⁷ cells/mL were achieved after 7 days of expansion at a specific growth rate (µ) of 0.61 ± 0.1 day⁻¹ with a perfusion rate (D) of 5.0 day⁻¹. A twofold increase in maximum cell density (to greater than 2.5 × 10⁷ cells/mL) was achieved when the medium dissolved oxygen was reduced (5% DO). Cell densities and viabilities >80% were maintained for extended production periods during which maintenance of pluripotency was confirmed by stable expression of pluripotency factors (SSEA-1 and Nanog), as well as the capacity to differentiate into all three germ layers. This work establishes a versatile biotechnological platform for the production of pluripotent cells and derivatives in an integrated, scalable and intensified stirred suspension culture. Copyright © 2012 Wiley Periodicals, Inc.

Citation

Ricardo P Baptista, David A Fluri, Peter W Zandstra. High density continuous production of murine pluripotent cells in an acoustic perfused bioreactor at different oxygen concentrations. Biotechnology and bioengineering. 2013 Feb;110(2):648-55